Fibrinogen is a clotting factor having a very important role in the final stage of the so-called blood-coagulation cascade. In the activation of, for example, the blood clotting system after a damage, soluble fibrinogen is converted, by thrombin, into insoluble fibrin which has an important role of contributing to the hemostasis and curing of a wound. Recently, a tissue adhesive was developed taking advantage of this theory in the final phase of the blood clotting, and it is now used as an adhesive in place of a suture of a soft organ such as liver or spleen or as a suture assistant in a surgical operation. A tissue adhesive is widely used also in other clinical situation.
For the application of the tissue adhesive to an affected part of the body, a "sequential method" wherein a fibrinogen solution and a thrombin solution are applied alternately to form a laminate coat or a "simultaneous method" wherein both the fibrinogen solution and thrombin solution are applied simultaneously to mix them together was mainly employed. However, the sequential method has a problem in that a major part of the fibrinogen solution flows down prior to the formation of a fibrin gel, and also the simultaneous method has a problem in that the formed fibrin gel is heterogeneous. Therefore, the effect of the tissue adhesive was limited in both the sequential method and simutaneous method.
Recently, a spray method has just started to be spread for solving the above-described problems. In this method, a fibrinogen solution and a thrombin solution in each syringe are simultaneously ejected together with a sterile gas to spray these two solutions in the form of a mist mixture. A preferred example of the devices for this method is described in International Patent Publication (PCT WO94-07420). However, even such a spray method has a problem to be solved for obtaining a complete effect of the tissue adhesive when a strong adhesion and closing effect are demanded in, for example, the treatment of great vessels and also the treatment of air leak in the lungs.
Investigations were made for improving the effect of the tissue adhesive by increasing the concentration of fibrinogen to find that, although the clotting strength and adhesive power of the fibrin gel obtained by the sequantial method are increased as the final concentration of fibrinogen in the mixture of the two components is increased, they are no more increased even when the final concentration of fibrinogen is increased to about 4% or higher [see "Kiso to Rinsho" (Basic and Clinical Report), 20, 2399 to 2405 (1986), and 23, 3735 to 3743 (1989)]. However, when the sequential method is employed, the fibrin gel obtained by such a test cannot be uniformly applied. Thus, it cannot be said that the advantage of the increase in the final concentration of fibrinogen has been put to practical use.
Japanese Patent Unexamined Published Application (hereinafter referred to as "J. P. KOKAI") No. Sho 61-293443 discloses a method for increasing the final concentration of fibrinogen in the mixture of the two components by changing the mixing ratio of the fibrinogen solution to the thrombin solution in its specification (in particular, in the "test results" in the detailed description of the Invention). In this specification, it is described that the limiting tensile strength depends on the final concentration of fibrinogen after mixing the two components.
However, in these tests, the adhesive is applied by so-called "mixing method" and, in addition, the final concentration of thrombin in the formation of the fibrin gel after mixing the two components is as low as 1 to 2 units/ml. Such a low thrombin concentration is far apart from the thrombin concentration (250 to 500 units/ml) actually employed in the clinical sites. It is known that particularly when the thrombin concentration is low, the clotting of fibrinogen necessitates a long period of time and, in addition, a sufficient hemostasis effect cannot be obtained in the closing treatment of the tissue. Although a test wherein the final concentration of thrombin after mixing the two components is as high as 500 units/ml is described therein, it is stated therein that the limiting tensile strength could not be determined. This fact suggests that the tests given in J. P. KOKAI No. Sho 61-293443 are unsuitable for the determination of the effects of the tissue adhesive to be used in a clinical site.
The main object of the invention disclosed in J. P. KOKAI No. Sho 61-293443 is to provide a device for preparing a fibrinogen solution from a freeze-dried preparation previously charged in a syringe, which is capable of shortening the dissolution time without reducing the limiting tensile strength of the adhesive. In this invention, the mixing ratio is varied so that a limiting tensile strength equal to that of an ordinary product (a concentration is 10% before the mixing) can be obtained even when the concentration of the fibrinogen solution is lowered. Therefore, the concentration described therein is not the practical one, and this invention does not provide any means for adhering a tissue which is suitable for the actual use for particularly obtaining a high effect of closing the wound.
Therefore, the object of the present invention is to provide a tissue adhesive suitable for spray application, which is to be used in the form of a two-pack preparation and which can be used at a high concentration and uniformly sprayed.
Another object of the present invention is to provide a tissue adhesive capable of uniformly forming fibrin of a high concentration to provide a high closing effect.
After intensive investigations made for the purpose of attaining the above-described objects, and on the basis of an idea that the spray method wherein the fibrinogen solution and thrombin solution can be homogeneously mixed together is the best method for the application of the tissue adhesive, the inventors have found the following fact: for improving the effect of fibrinogen by increasing the final concentration of fibrinogen in the mixture of the two components, it is effective that the concentration of fibrinogen which can be sprayed is made as high as possible and also that the mixing ratio of the fibrinogen solution and thrombin solution is changed. Further, the structure of a device for conducting this method has been made clear, and the present invention capable of providing the tissue adhesive suitable for use in a method wherein a remarkable effect is obtained has been completed.